Synthetic composition for treating metabolic disorders

ABSTRACT

A method for treating metabolic disorders includes determining a treatment group comprising obese non-infant humans; formulating a composition comprising one or more synthetic non-fucosylated human milk oligosaccharides (HMOs) selected from lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT) and/or 2′-fucosyllactose (2′-FL), 3-fucosyllactose (3-FL), difucosyllactose (DFL), and lacto-N-fucopentaose I (LNFP-I), that are effective for: increasing in the gastrointestinal microbiota of a non-infant human during a treatment period, the relative abundance of  Bifidobacterium adolescentis  and reducing a precursor condition for a metabolic disorder associated with development of one or more of obesity-induced pre-diabetes and type 2 diabetes, the precursor condition selected from gut permeability, metabolic endotoxemia, low-grade metabolic inflammation, and body fat percentage; and reducing the precursor condition in at least one non-infant human in the treatment group by providing the composition to the at least one non-infant human during the treatment period.

CROSS-REFERENCES TO RELATED APPLICATIONS

The present application is a Continuation of U.S. application Ser. No.17/093,337 filed Nov. 9, 2020. U.S. application Ser. No. 17/093,337 is aContinuation of and claims priority to U.S. application Ser. No.15/104,794 filed Jun. 15, 2016, now patented U.S. Pat. No. 10,828,313,which is the U.S. National Stage Entry of PCT/DK2015/050385 which claimspriority to DK application PA 2014 70768, the entire contents of each ofthe aforementioned applications are incorporated herein by reference forall purposes permissible under relevant patent laws and rules.

FIELD

This disclosure relates generally to compositions and methods for thetreatment of metabolic disorders such as obesity and obesity inducedpre-diabetes and type 2 diabetes.

BACKGROUND

Diabetes type 2 is a metabolic disorder that is characterized byhyperglycaemia due to insulin resistance and relative lack of insulinand that is a rapidly growing global epidemic. The InternationalDiabetes Federation (IDF) reports that as of 2013 there were more than382 million people living with diabetes, and a further 316 million withimpaired glucose tolerance who are at high risk from the disease (IDFDiabetes Atlas, 6^(th) edn.). The World Health Organization (WHO)furthermore estimates that 90 percent of people around the world whosuffer from diabetes suffer from type 2 diabetes. Long-termcomplications from high blood sugar can include heart disease, strokes,diabetic retinopathy, kidney failure, and poor blood flow in the limbs.

Since it is unlikely that there has been a dramatic alteration ingenetic factors in the past decades, environmental factors must play akey role in the rapid rise in diabetes. Gut microbiota may be a keyfactor with populations showing marked differences between healthy,obese, and type 2 diabetic patients. The dysbiosis of gut microbiota hasthe potential to affect host metabolism and energy storage and to affectgut permeability and, as a consequence, give rise to metabolicendotoxemia and higher plasma lipopolysaccharide (LPS). In addition, gutpeptides such as glucagon-like peptide 1 (GLP1) and GLP2 can play keyroles in these processes. For example, GLP2, which is secreted byintestine L cells, is a key regulator of intestinal permeability.Therapeutic regimes that target intestinal microbiota and intestinalbarrier therefore show a broad prospect in treating diabetes.

Recent insights suggest that an altered composition and diversity of gutmicrobiota could play an important role in the development of metabolicdisorders such as obesity and diabetes. Gut microbiota does not onlyparticipate in whole-body metabolism by affecting energy balance andglucose metabolism, but is also involved in development of the low-gradeinflammation, associated with obesity and related metabolic disorderssuch as diabetes. The association between inflammation and type 2diabetes is seen in epidemiological studies indicating a rise inacute-phase response proteins in serum of type 2 diabetic patientscompared with controls. Later, a specific link between inflammatory andmetabolic responses was made with the discovery that compared with leantissue, obese adipose tissue secretes inflammatory cytokines and thatthese inflammatory cytokines themselves can inhibit insulin signalling.The definitive proof of a connection between inflammatory mediators andinsulin resistance in obesity and type 2 diabetes came from geneticstudies that interfered with inflammatory mediators and demonstratedbeneficial effects of this interference on insulin action.

In recent years, gut microbiota derived LPS has been shown to beinvolved in the onset and progression of inflammation, and inpathological situations, such as obesity and type 2 diabetes, LPS play amajor role in the onset of disease. After only one week of a high-fatdiet in mice, commensal intestinal bacteria are translocated from theintestine into adipose tissue and the blood where they can induceinflammation. This metabolic bacteraemia is characterized by anincreased co-localization with dendritic cells from the intestinallamina propria and by an augmented intestinal mucosal adherence ofnon-pathogenic Escherichia coli. The bacterial translocation processfrom intestine towards tissue with resulting inflammation was reversedby six weeks of treatment with the probiotic strain Bifidobacteriumanimalis subsp. lactis 420, suggesting an involvement of the microbiota.

EP-A-1332759 discloses that oral doses of 2′-FL, 3′-SL, 6′-SL, LNnT andsialic acid promote insulin secretion in type 2 diabetes-model mice.

EP-A-2143341 discloses that a mixture of GOS, sialylatedoligosaccharides and N-acylated oligosaccharides reduces triglycerideconcentration in liver in model mice.

EP-A-2332552 discloses that 3′-SL and 6′-SL reduce/prevent fataccumulation in the liver and other organs in high-fat diet mice andrats.

WO 2013/057061 discloses a composition for increasing insulinsensitivity and/or reducing insulin resistance. The composition containslong chain polyunsaturated fatty acids, probiotics and a mixture ofoligosaccharides containing at least one of lacto-N-neotetraose (LNnT)and lacto-N-tetraose (LNT), at least one N-acetylated oligosaccharidedifferent from LNnT and LNT, at least one sialylated oligosaccharide andat least one neutral oligosaccharide, for use in increasing insulinsensitivity and/or reducing insulin resistance. This composition canalso contain 2′-O-fucosyllactose (2′-FL). The composition isparticularly adapted for use in infants who were born preterm and/or whoexperienced IUGR, and in pregnant women suffering from gestationaldiabetes. It is also stated that the composition can be given tochildren, adolescents, and adults suffering from insulin resistanceand/or type II diabetes. It is stated that the efficacy of thecomposition can be the result of the synergistic combination of immunitymodulator effects triggered by the probiotics and the LC-PUFA throughtheir stimulation with the specific oligosaccharide mixture.

Most current therapeutic approaches aim at treating the consequencesrather than causes of the impaired metabolism. This strategy is notefficient and therefore, there has remained a need for therapies thatreduce intestinal permeability, endotoxemia and low-grade inflammationin patients with metabolic disorders to improve glucose and insulinsensitivity, and which are safe with little or no adverse side effects.

SUMMARY

A method is provided for treating metabolic disorders. In a firstaspect, the method includes: determining a treatment group comprisingobese non-infant humans, formulating a composition comprising aneffective amount of one or more synthetic fucosylated human milkoligosaccharides (HMO) selected from 2′-fucosyllactose (2′-FL),3-fucosyllactose (3-FL), difucosyllactose (DFL), andlacto-N-fucopentaose I (LNFP-I), that are effective for increasing inthe gastrointestinal microbiota of the non-infant human during atreatment period, the relative abundance of Bifidobacterium adolescentisand reducing a precursor condition for a metabolic disorder associatedwith development of one or more of obesity-induced pre-diabetes and type2 diabetes, the precursor condition selected from gut permeability,metabolic endotoxemia, low-grade metabolic inflammation, and body fatpercentage. The method further includes reducing the precursor conditionin at least one non-infant human in the treatment group by providing thecomposition to the at least one non-infant human during the treatmentperiod.

In a second aspect, the method includes: determining a treatment groupcomprising obese non-infant humans, formulating a composition comprisingan effective amount of one or more synthetic non-fucosylated human milkoligosaccharides (HMOs) selected from lacto-N-tetraose (LNT),lacto-N-neotetraose (LNnT), that are effective for: increasing in thegastrointestinal microbiota of the non-infant human during a treatmentperiod, the relative abundance of Bifidobacterium adolescentis, andreducing a precursor condition for a metabolic disorder associated withdevelopment of one or more of obesity-induced pre-diabetes and type 2diabetes, the precursor condition selected from gut permeability,metabolic endotoxemia, low-grade metabolic inflammation, and body fatpercentage. The method further includes reducing the precursor conditionin at least one non-infant human in the treatment group by providing thecomposition to the at least one non-infant human during the treatmentperiod.

According to a third aspect of the disclosure, the method includes:determining a treatment group comprising obese non-infant humans, fformulating a composition comprising an effective amount of two or moresynthetic neutral human milk oligosaccharides (HMOs) selected from2′-fucosyllactose (2′FL), 3-fucosyllactose (3-FL), difucosyllactose(DFL), lacto-N-fucopentaose I (LNFP-I), lacto-N-tetraose (LNT), andlacto-N-neotetraose (LNnT), wherein the selected HMOs are effective for:increasing in the gastrointestinal microbiota of the non-infant humanduring a treatment period, the relative abundance of Bifidobacteriumadolescentis, and reducing a precursor condition for a metabolicdisorder associated with development of one or more of obesity-inducedpre-diabetes and type 2 diabetes, the precursor condition selected fromgut permeability, metabolic endotoxemia, low-grade metabolicinflammation, and body fat percentage. The method further includesreducing the precursor condition in at least one non-infant human in thetreatment group by providing the composition to the at least onenon-infant human during the treatment period.

In some embodiments, the precursor condition for the metabolic disorderassociated with development of the one or more of obesity-inducedpre-diabetes and type 2 diabetes is gut permeability. In certainembodiments, the reduced precursor condition is body fat percentage.

In various embodiments, the method includes increasing, in thegastrointestinal tract of the non-infant human, a level of glucagon-likepeptide selected from GLP-1 and/or GLP-2 relative to the level of theselected glucagon-like peptide prior to the treatment period.

In certain embodiments, the treatment period includes an initialtreatment phase and a maintenance phase. The effective amount of theselected HMO is from about 2.5 g to about 7.5 g daily during the initialphase, and the effective amount of the selected HMO is from about 1 g toabout 2.5 g daily during the maintenance phase.

In some embodiments, a single synthetic HMO of the selected HMOs isadministered in a unit dosage form. In certain embodiments, the obesenon-infant human is a prepubescent child.

DETAILED DESCRIPTION OF THE INVENTION

In one aspect, various embodiments of this invention provide a syntheticcomposition for use in one or more of the following: reducing intestinalpermeability, reducing endotoxemia, reducing low-grade inflammation,reducing body fat percentage,

increasing the abundance of bifidobacteria, and/or increasing the levelsof the gut hormones GLP-1 and GLP-2, in a patient having a metabolicdisorder, for example obesity, obesity induced pre-diabetes and obesityinduced type 2 diabetes, characterized in that the synthetic compositioncontains an effective amount of one or more human milk monosaccharidesor one or more human milk oligosaccharides (“HMOs”) or both. Thesynthetic composition is preferably a nutritional composition. Thecomposition can further comprise a source of threonine, serine and/orproline. Preferably the human milk oligosaccharides include bothfucosylated and core HMOs such as LNT and LNnT HMOs. The human milkoligosaccharides can also include sialylated HMOs. Alternatively, thehuman milk oligosaccharides include both fucosylated and sialylated HMOsand the human milk oligosaccharides can also include backbone HMOs suchas LNT and LNnT. The patient can be a paediatric or adult patient,preferably a prepubescent child.

In another aspect, this disclosure provides a method for one or more ofthe following: reducing intestinal permeability, reducing endotoxemia,reducing low-grade inflammation, reducing body fat percentage,increasing the abundance of bifidobacteria, and/or increasing the levelsof the gut hormones GLP-1 and GLP-2, in a patient having a metabolicdisorder, for example obesity, obesity induced pre-diabetes and obesityinduced type 2 diabetes, the method comprising orally administering tothe patient an effective amount of one or human milk monosaccharides orone or more human milk oligosaccharides, or both, preferably in the formof a synthetic composition. The patient can be a paediatric or adultpatient, preferably a prepubescent child.

In a further aspect, this disclosure relates to a use of one or morehuman milk monosaccharides or one or more human milk oligosaccharides orboth, preferably in the form of a synthetic composition, for one or moreof the following: reducing intestinal permeability, reducing endotoxemiain a patient, reducing low-grade inflammation, reducing body fatpercentage, increasing the abundance of bifidobacteria, and/orincreasing the levels of the gut hormones GLP-1 and GLP-2, in a patienthaving a metabolic disorder, for example obesity, obesity inducedpre-diabetes and obesity induced type 2 diabetes. The patient can be apaediatric or adult patient, preferably a prepubescent child.

It has been surprisingly found that human milk monosaccharides,advantageously sialic acid and/or fucose, and human milkoligosaccharides, advantageously 2′-FL, 3-FL, LNT, LNnT, 3′-SL, 6′-SL,DFL, DSLNT and/or LNFP-I, not only modulate inflammation and microbiotain the GI tract, but also decrease gut permeability, reduce endotoxemiaand low-grade inflammation, improve body composition (by reducing bodyfat percentage), increase the abundance of bifidobacteria, and increasethe levels of the gut hormones GLP-1 and GLP-2 in human patients.Preferably the abundance of Bifidobacteria adolescentis is increased.This can result in lower chronic inflammation, improved insulinsensitivity and reduced insulin resistance. Obese and pre-diabeticpatients can be stabilized and the progression to diabetes slowed,stopped or reversed. Diabetic patients can be stabilized or at least theprogression to diabetes with complications slowed.

In certain aspects, the synthetic composition preferably contains one ormore core HMOs and/or one or more fucosylated HMOs, more preferably oneor more core HMOs and one or more fucosylated HMOs. Even more preferablythe core HMO is selected from the group consisting of LNT, LNnT, LNH,LNnH and pLNnH, particularly LNT and LNnT, and the fucosylated HMO isselected from the group consisting of 2′-FL, 3-FL, DFL and LNFP-I,particularly 2′-FL. Advantageously, the synthetic composition contains2′-FL and LNT and/or LNnT.

The term “patient” preferably means a human that can be a paediatric oradult patient. However, a “patient” can also be any other mammal.

The term “oral administration” preferably means any conventional formfor the oral delivery of a composition to a patient that causes thedeposition of the composition in the gastrointestinal tract (includingthe stomach) of the patient. Accordingly, oral administration includesswallowing of composition by the patient, enteral feeding through anaso-gastric tube, and the like.

The term “effective amount” preferably means an amount of a compositionthat provides a human milk monosaccharide or human milk oligosaccharidein a sufficient amount to render a desired treatment outcome in apatient. An effective amount can be administered in one or more doses tothe patient to achieve the desired treatment outcome.

The term “human milk monosaccharide” or “HMS” preferably means amonosaccharide found in human breast milk. Examples include sialic acidand L-fucose. In human milk, the sialic acid is N-acetylneuraminic acid.

The term “human milk oligosaccharide” or “HMO” preferably means acomplex carbohydrate found in human breast milk that can be in acidic orneutral form. More than about 200 different HMO structures are known toexist in human breast milk (Urashima et al.: Milk Oligosaccharides, NovaBiomedical Books, New York, 2011). HMOs can be core, fucosylated andsialylated oligosaccharides. Core HMOs consist of Glu, Gal and GlcNAcand are devoid of Fuc and sialic acid. Examples of core HMOs includelacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT), lacto-N-neohexaose(LNnH), lacto-N-hexaose (LNH) and p-lacto-N-neohexaose (pLNnH). FucosylHMOs are fucosylated lactoses or fucosylated core HMOs such as2′-fucosyllactose (2′-FL), lacto-N-fucopentaose I (LNFP-I),lacto-N-difucohexaose I (LNDFH-I), 3-fucosyllactose (3-FL),difucosyllactose (DFL), lacto-N-fucopentaose III (LNFP-III),fucosyl-para-lacto-N-neohexaose (F-pLNnH), lacto-N-difucohexaose I(LNDFH-I), fucosyl-lacto-N-hexaose II (FLNH-II), lacto-N-fucopentaose V(LNFP-V), lacto-N-difucohexaose II (LNDFH-II), fucosyl-lacto-N-hexaose I(FLNH-I), fucosyl-lacto-N-hexaose III (FLNH-III) andfucosyl-para-lacto-N-neohexaose (F-pLNnH). Sialyl HMOs are sialylatedlactoses or sialylated core HMOs such as 3′,6-disialyllacto-N-tetraose(DSLNT), 6′-sialyllactose (6′-SL), 3′-sialyllactose (3′-SL),6′-sialyllacto-N-neotetraose (LST c), 3′-sialyllacto-N-tetraose (LST a)and 6-sialyllacto-N-tetraose (LST b). HMOs containing both sialyl andfucosyl groups may be considered to belong to either of the latter twogroups. Examples for sialylated and fucosylated HMOs includedisialyl-fucosyl-lacto-N-hexaose II (DSFLNH-II),fucosyl-sialyl-lacto-N-neohexaose I (FSLNnH-I),fucosyl-sialyl-lacto-N-hexaose I (FSLNH-I) and3-fucosyl-3′-sialyllactose (FSL).

The term “intestinal permeability” preferably means the permeability ofthe intestinal mucosa of a patient, permitting the absorption of vitalnutrients from the gut lumen while presenting a barrier against thepassage of pathogenic substances into the patient's body.

The term “endotoxemia” preferably means the presence of endotoxins, suchas gut microbiota-derived lipopolysaccharides (LPS) in the blood of apatient.

The term “low-grade inflammation” preferably means an immune systemresponse of a patient characterized by altered levels ofpro-inflammatory and anti-inflammatory cytokines as well as numerousother markers of immune system activity in response to an injuriousstimuli.

Body fat percentage preferably means total mass of body fat divided bytotal mass of the body.

The HMOs can be isolated or enriched by well-known processes frommilk(s) secreted by mammals including, but not limited to human, bovine,ovine, porcine, or caprine species. The HMOs can also be produced bywell-known processes using microbial fermentation, enzymatic processes,chemical synthesis, or combinations of these technologies. As examples,using chemistry LNnT can be made as described in WO 2011/100980 and WO2013/044928, LNT can be synthesized as described in WO 2012/155916 andWO 2013/044928, a mixture of LNT and LNnT can be made as described in WO2013/091660, 2′-FL can be made as described in WO 2010/115934 and WO2010/115935, 3-FL can be made as described in WO 2013/139344, 6′-SL andsalts thereof can be made as described in WO 2010/100979, sialylatedoligosaccharides can be made as described in WO 2012/113404 and mixturesof human milk oligosaccharides can be made as described in WO2012/113405. As examples of enzymatic production, sialylatedoligosaccharides can be made as described in WO 2012/007588, fucosylatedoligosaccharides can be made as described in WO 2012/127410, andadvantageously diversified blends of human milk oligosaccharides can bemade as described in WO 2012/156897 and WO 2012/156898. With regard tobiotechnological methods, WO 01/04341 and WO 2007/101862 describe how tomake core human milk oligosaccharides optionally substituted by fucoseor sialic acid using genetically modified E. coli.

The human milk monosaccharides and/or oligosaccharides can be in theform of one or more core HMOs and one or more fucosylated HMOs.Alternatively, one or more core HMOs and one or more sialylated HMOs canbe used. In a further alternative, one or more fucosylated HMOs and oneor more sialylated HMOs can be used. In a preferred embodiment, one ormore core HMOs, one or more sialylated HMOs and one or more fucosylatedHMOs are used.

The term “glycaemic index” or “GI” is defined as the incremental areaunder the two-hour blood glucose response curve (AUC) following a12-hour fast and ingestion of a food with a certain quantity ofavailable carbohydrate (usually 50 g). The AUC of the test food isdivided by the AUC of the standard (glucose, the standard, has a GI of100) and multiplied by 100. The average GI value is calculated from datacollected in 10 human subjects. Both the standard and test food mustcontain an equal amount of available carbohydrate. The result gives arelative ranking for each tested food. Tables reporting commonlyaccepted GI values for a variety of foods are available including theinternational GI database maintained by the University of Sydney, andavailable on the internet at: www.glycemicindex.com.

The synthetic composition can take any suitable form. For example, thecomposition can be in the form of a nutritional composition whichcontains other macronutrients such as proteins, lipids or othercarbohydrates. The synthetic composition can also be a pharmaceuticalcomposition.

Nutritional Compositions

A nutritional composition can contain sources of protein, lipids and/ordigestible carbohydrates and can be in solid, powdered or liquid forms.The composition can be designed to be the sole source of nutrition or anutritional supplement.

Suitable protein sources include intact, hydrolysed, and partiallyhydrolysed protein, which can be derived from any suitable source suchas milk (e.g., casein, whey), animal (e.g., meat, fish), cereal (e.g.,rice, corn), and vegetable (e.g., soy, potato, pea), insect (e.g.,locust) and combinations of these sources. Examples of the source ofprotein include whey protein concentrates, whey protein isolates, wheyprotein hydrolysates, acid caseins, sodium casemates, calcium casemates,potassium casemates, casein hydrolysates, milk protein concentrates,milk protein isolates, milk protein hydrolysates, non-fat dry milk,condensed skim milk, soy protein concentrates, soy protein isolates, soyprotein hydrolysates, pea protein concentrates, pea protein isolates,pea protein hydrolysates, collagen proteins, and combinations of thesesources.

The amount of protein is preferably sufficient to provide about 5 toabout 30% of the energy of the nutritional composition; for exampleabout 10% to about 25% of the energy. Within these ranges, the amount ofprotein can vary depending upon the nutritional needs of the intendedindividual.

The nutritional compositions can also include free amino acids such astryptophan, glutamine, tyrosine, methionine, cysteine, taurine,arginine, carnitine, threonine, serine and proline and combinations ofthese amino acids. Threonine, serine and proline are important aminoacids for the production of mucin which aids gut barrier function.

Any suitable source of other carbohydrates can be used. Examples includemaltodextrin, hydrolyzed or modified starch or corn starch, glucosepolymers, corn syrup, corn syrup solids, rice-derived carbohydrates,sucrose, glucose, fructose, lactose, high fructose corn syrup, honey,sugar alcohols (e.g., maltitol, erythritol, sorbitol, etc.),isomaltulose, sucromalt, pullulan, potato starch, slowly-digestedcarbohydrates, dietary fibres such as oat fibre, soy fibre, gum arabic,sodium carboxymethylcellulose, methylcellulose, guar gum, gellan gum,locust bean gum, konjac flour, hydroxypropyl methylcellulose, tragacanthgum, karaya gum, gum acacia, chitosan, arabinogalactans, glucomannan,xanthan gum, alginate, pectin, low and high methoxy pectin, cerealbeta-glucans (i.e., oat beta-glucan, barley beta-glucan), carrageenanand psyllium, Fibersol™ other resistant starches, and combinations ofthese carbohydrate.

Preferably the carbohydrate source includes low glycaemic indexcarbohydrates having a GI score of 55 or below. Examples of lowglycaemic index carbohydrates include sucromalt, Fibersol™ (inulin),maltodextrins having a dextrose equivalence (DE) of less than 15, ricesyrup having a dextrose equivalence of less than 15,fructooligosaccharides, resistant starches, starches, fruit sourcedfibres, vegetable sourced fibres, whole grains, beta-glucans, soyfibres, oat fibres, locust bean gum, konjac flour, hydroxypropylmethylcellulose, gum acacia, chitosan, arabinogalactans, xanthan gum,alginate, low and high methoxy pectin, carrageenan, psyllium,isomaltulose, glycerine and sugar alcohols.

The nutritional compositions can include carbohydrates in an amountsufficient to provide about 30 to about 70% of the energy of thecomposition, for example about 35 to about 65% of the energy. Withinthese parameters, the amount of carbohydrate can vary widely.

Suitable lipid sources include coconut oil, fractionated coconut oil,soy oil, corn oil, olive oil, safflower oil, high oleic safflower oil,medium chain triglycerides, sunflower oil, high oleic sunflower oil,palm and palm kernel oils, palm olein, canola oil, marine oils,cottonseed oils and combinations of these oils. Fractionated coconutoils are a suitable source of medium chain triglycerides. The lipids cancontain polyunsaturated fatty acids such as n-3 LC-PUFA. The n-3 LC-PUFAcan be a C20 or a C22 n-3 fatty acid. Preferably the n-3 LC-PUFA isdocosahexanoic acid (DHA, C22:6, n-3). The source of LC-PUFA can be, forexample, egg lipids, fungal oil, low EPA fish oil or algal oil.

The nutritional compositions can include lipids in an amount sufficientto provide about 10 to about 50% of energy of the nutritionalcomposition, for example about 15 to about 40% of the energy.

The nutritional composition preferably also includes vitamins andminerals. If the nutritional composition is intended to be a sole sourceof nutrition, it preferably includes a complete vitamin and mineralprofile. Examples of vitamins include vitamins A, B-complex (such as B1,B2, B6 and B12), C, D, E and K, niacin and acid vitamins such aspantothenic acid, folic acid and biotin. Examples of minerals includecalcium, iron, zinc, magnesium, iodine, copper, phosphorus, manganese,potassium, chromium, molybdenum, selenium, nickel, tin, silicon,vanadium and boron.

The nutritional composition can also include a carotenoid such aslutein, lycopene, zeaxanthin, and beta-carotene. The total amount ofcarotenoid included can vary from about 0.001 μg/ml to about 10 μg/ml.Lutein can be included in an amount of from about 0.001 μg/ml to about10 μg/ml, preferably from about 0.044 μg/ml to about 5 g/ml of lutein.Lycopene can be included in an amount from about 0.001 μg/ml to about 10μg/ml, preferably about 0.0185 mg/ml to about 5 g/ml of lycopene.Beta-carotene can comprise from about 0.001 μg/ml to about 10 mg/ml, forexample about 0.034 μg/ml to about 5 μg/ml of beta-carotene. Thenutritional composition can also include a source of anthocyanidins.This can be in the form of a fruit or a fruit extract. Particularlyuseful fruits and fruit extracts include plum/prune, apple, pear,strawberry, blueberry, raspberry, cherry, and their combinations.

The nutritional composition can also contain various other conventionalingredients such as preservatives, emulsifying agents, thickeningagents, buffers, fibres and prebiotics (e.g. fructooligosaccharides,galactooligosaccharides), probiotics (e.g. B. animalis subsp. lactisBB-12, B. lactis HN019, B. lactis Bi07, B. infantis ATCC 15697, L.rhamnosus GG, L. rhamnosus HNOOI, L. acidophilus LA-5, L. acidophilusNCFM, L. fermentum CECT5716, B. longum BB536, B. longum AH1205, B.longum AH1206, B. breve M-16V, L. reuteri ATCC 55730, L. reuteri ATCCPTA-6485, L. reuteri DSM 17938), antioxidant/anti-inflammatory compoundsincluding tocopherols, carotenoids, ascorbate/vitamin C, ascorbylpalmitate, polyphenols, glutathione, and superoxide dismutase (melon),other bioactive factors (e.g. growth hormones, cytokines, TFGβ),colorants, flavours, and stabilizers, lubricants, and so forth.

The nutritional composition can be in the form of a food, solublepowder, a liquid concentrate, or a ready-to-use formulation. Thecomposition can be eaten, drunk or can be fed via a nasogastric. Variousflavours, fibres, and other additives can also be present.

The nutritional compositions can be prepared by any commonly usedmanufacturing techniques for preparing nutritional compositions in solidor liquid form. For example, the composition can be prepared bycombining various feed solutions. A protein-in-fat feed solution can beprepared by heating and mixing the lipid source and then adding anemulsifier (e.g. lecithin), fat soluble vitamins, and at least a portionof the protein source while heating and stirring. A carbohydrate feedsolution is then prepared by adding minerals, trace and ultra traceminerals, thickening or suspending agents to water while heating andstirring. The resulting solution is held for 10 minutes with continuedheat and agitation before adding carbohydrates (e.g., the HMOs anddigestible carbohydrate sources). The resulting feed solutions are thenblended together while heating and agitating and the pH adjusted to6.6-7.0, after which the composition is subjected to high-temperatureshort-time processing during which the composition is heat treated,emulsified and homogenized, and then allowed to cool. Water solublevitamins and ascorbic acid are added, the pH is adjusted to the desiredrange if necessary, flavours are added, and water is added to achievethe desired total solid level.

For a liquid product, the resulting solution can then be asepticallypacked to form an aseptically packaged nutritional composition. In thisform, the nutritional composition can be in ready-to-feed orconcentrated liquid form. Alternatively, the composition can bespray-dried and processed and packaged as a reconstitutable powder.

The nutritional composition can also be in the form of a food such as anutritional bar, a yoghurt, etc. These forms can be produced usingstandard technologies and processes.

When the nutritional product is a ready-to-feed nutritional liquid, thetotal concentration of HMSs/HMOs in the liquid, by weight of the liquid,is from about 0.0001% to about 2.0%, including from about 0.001% toabout 1.5%, including from about 0.01% to about 1.0%. When thenutritional product is a concentrated nutritional liquid, the totalconcentration of HMSs/HMOs in the liquid, by weight of the liquid, isfrom about 0.0002% to about 4.0%, including from about 0.002% to about3.0%, including from about 0.02% to about 2.0%.

Unit Dosage Forms

The synthetic composition of this disclosure can also be in a unitdosage form such as a capsule, tablet or sachet. For example, thecomposition can be in a tablet form comprising the human milkmonosaccharides and/or oligosaccharides, and one or more additionalcomponents to aid formulation and administration, such as diluents,excipients, antioxidants, lubricants, colorants, binders, disintegrants,and the like.

Suitable diluents, excipients, lubricants, colorants, binders, anddisintegrants include polyethylene, polyvinyl chloride, ethyl cellulose,acrylate polymers and their copolymers, hydroxyethyl-cellulose,hydroxypropylmethyl-cellulose (HPMC), sodium carboxymethylcellulose,polyhydroxyethyl methacrylate (PHEMA), polyvinyl alcohol (PVA),polyvinyl pyrrolidone (PVP), polyethylene oxide (PEO), or polyacrylamide(PA), carrageenan, sodium alginate, polycarbophil, polyacrylic acid,tragacanth, methyl cellulose, pectin, natural gums, xanthan gum, guargum, karaya gum, hypromellose, magnesium stearate, microcrystallinecellulose, and colloidal silicon dioxide. Suitable antioxidants arevitamin A, carotenoids, vitamin C, vitamin E, selenium, flavonoids,polyphenols, lycopene, lutein, lignan, coenzyme Q10 (“CoQIO”) andglutathione. The unit dosage forms, especially those in sachet form, canalso include various nutrients including macronutrients.

Administration Dosing

For reducing intestinal permeability, endotoxemia, low-gradeinflammation and/or body fat percentage, and/or increasing the abundanceof bifidobacteria and/or the levels of the gut hormones GLP-1 and GLP-2in a person, the amount of human milk mono and/or oligosaccharide(s)required to be administered to the person will vary depending uponfactors such as the risk and condition severity, the age of the person,the form of the composition, and other medications being administered tothe person. However, the required amount can be readily set by a medicalpractitioner and would generally be in the range from about 10 mg toabout 20 g per day, in certain embodiments from about 10 mg to about 15g per day, from about 100 mg to about 10 g per day, in certainembodiments from about 500 mg to about 10 g per day, in certainembodiments from about 1 g to about 7.5 g per day. An appropriate dosecan be determined based on several factors, including, for example, thebody weight and/or condition of the patient being treated, the severityof the condition, being treated, other ailments and/or diseases of theperson, the incidence and/or severity of side effects and the manner ofadministration. Appropriate dose ranges can be determined by methodsknown to those skilled in the art. During an initial treatment phase,the dosing can be higher (for example 200 mg to 20 g per day, preferably500 mg to 15 g per day, more preferably 1 g to 10 g per day, in certainembodiments 2.5 g to 7.5 g per day). During a maintenance phase, thedosing can be reduced (for example, 10 mg to 10 g per day, preferably100 mg to 7.5 g per day, more preferably 500 mg to 5 g per day, incertain embodiments 1 g to 2.5 g per day).

A synthetic composition of this disclosure can be co-administered to apatient who is also receiving a standard-of-care medication for obesityor diabetes.

EXAMPLES

Examples are now described to further illustrate the features of thedisclosed methods:

Example 1—Treating High Fat Diet Induced Obesity and Diabetes

10-week-old C57BL/6J mice (120 mice) are housed in groups of five miceper cage, with free access to food and water. The mice are divided into12 groups of 10 mice, one control group and 11 treatment groups. All ofthe mice are fed a high-fat (HF) diet (60% fat and 20% carbohydrates[kcal/100 g], or an HF diet supplemented with HMS/HMO (20 g/kg of diet)for 8 weeks. Food and water intake are recorded twice a week. The 11treatment groups are each administered one of the following: a) sialicacid, b) L-fucose, c) 2′-FL, d) 3-FL, e) 3′-SL, f) 6′-SL, g) LNT, h)LNnT, i) LNFP-I, j) DSLNT and k) a combination of these saccharides. Thecontrol group is administered the HF diet only. Fresh food is givendaily.

Intraperitoneal or oral glucose tolerance tests are performed asfollows: 6-h-fasted mice are injected with glucose into the peritonealcavity (1 g/kg glucose, 20% glucose solution) or by gavage (3 g/kgglucose, 66% glucose solution). Blood glucose is determined with aglucose meter (Roche Diagnostics) on 3.5 μl blood collected from the tipof the tail vein. A total of 20 μl blood is sampled 30 min before and 15or 30 min after the glucose load to assess plasma insulin concentration.

To assess intestinal permeability in vivo, the intestinal permeabilityof 4000 Da fluorescent dextran-FITC (DX-4000-FITC) is measured. Mice arefasted for 6 h before given DX-44-FITC by gavage (500 mg/kg body weight,125 mg/ml). After 1 h and 4 h, 120 ml of blood is collected from the tipof the tail vein. The blood is centrifuged at 4° C., 12 000 g for 3 min.Plasma is diluted in an equal volume of PBS (pH 7.4) and analysed forDX-4000-FITC concentration with a fluorescence spectrophotometer at anexcitation wavelength of 485 nm and emission wavelength of 535 nm.Standard curves are obtained by diluting FITC-dextran in non-treatedplasma diluted with PBS (1:3 v/v).

Plasma LPS, cytokines and gut hormones are determined as follows. PlasmaLPS concentration is measured using a kit based upon a Limulusamoebocyte extract (LAL kit endpoint-QCL1000). Samples are diluted 1/40to 1/100 and heated for 20 cycles of 10 min at 68° C. and 10 min at 4°C. An internal control for LPS recovery is included in the calculation.Plasma cytokines (interleukin (IL) 1α, IL1β, tumour necrosis factor(TNF)α, IL6, monocyte chemoattractant protein (MCP)-1, macrophageinflammatory protein (MIP)-1α, IL10, interferon (INF) c, IL15, IL18) andgut hormones (GLP-1 (active), GIP (total), amylin (active), pancreaticpolypeptide) are respectively determined in duplicate by using aBio-Plex Multiplex kit, or a mouse gut hormones panel (LincoPlex), andmeasured by using Luminex technology, an EIA kit (GLP-2 EIA kit) is usedto quantify GLP-2.

Mice are anaesthetised (ketamine/xylazine, intraperineally, 100 and 10mg/kg, respectively) after a 5 h period of fasting, and blood samplesand tissues are harvested for further analysis. Mice are killed bycervical dislocation. Liver, caecum (full and empty), muscles (vastuslateralis), and adipose tissues (mesenteric and corresponding lymphnodes, epididymal, subcutaneous and visceral) are precisely dissectedand weighed. The intestinal segments (jejunum, colon) are immersed inliquid nitrogen, and stored at −80° C., for further analysis.

To assess the microbiota profile, the caecal contents collected postmortem from mice are stored at −80° C. DNA is isolated from the caecalcontent samples using QIAamp DNA Stool Mini Kit. The DNA concentrationof extracts is measured using NanoDrop. Aliquots of 100 ng of extractedDNA are subjected to PCR using the 16S rDNA universal heteroduplexanalysis (HDA) primers HDA1-GC and HDA2 which are disclosed in Walter etal. Appl. Environ. Microbiol. 66, 297 (2000)) at 56° C. for strandannealing. Initial denaturation at 94° C. for 4 min is followed bythirty cycles of 30 s at 94° C., 30 s at 56° C. and 1 min at 72° C. Thequality of PCR products is verified by agarose gel electrophoresis.

Amplified 16S rDNA fragments are separated by denaturing gradient gelelectrophoresis (DGGE) using an INGENYphorU system equipped with 6%polyacrylamide gels with a denaturant in the range of 30-55%, where 100%denaturant is equivalent to 7M-urea and 40% formamide. Electrophoresisis carried out at 130 V for 4-5 hours at 60° C. Polyacrylamide gels arestained with GelRede nucleic acid stain for 45 min, destained inultrapure water and viewed under UV light. Bands of interest are excisedfrom gels and lysed in ultrapure water. Extracted DNA is re-amplifiedusing the same primers and PCR conditions.

To purify the bacterial DNA, PCR products are reloaded on a denaturantgradient gel followed by excision and lysis of selected bands. DNAsamples recovered from lysed bands of the second DGGE are re-amplifiedby PCR before purification using the QIAquick PCR Purification Kit andsequenced. Species identification is done using the Ribosomal MicrobiomeDatabase Project Classifier tool. Because of the limited sensitivity ofDGGE to quantify microbial diversity, the microbial composition of DNAsamples is also analysed using high-throughput sequencing.

The V5-V6 region of 16S rRNA from caecal content DNA samples isamplified using a forward primer and a reverse primer which are bothdisclosed in Andersson et al. PloS ONE 3, e2836 (2008)). Amplicons arepyrosequenced using a Roche 454 GS-FLX system. Sequences of at least 240nucleotides and containing no more than two undetermined bases areretained for taxonomic assignment. The QIIME software is used forchimera check and the Greengenes database is used for classification.Bacterial diversity is determined at the phylum, family and genuslevels.

To assess bacterial translocation from intestine into tissues,mesenteric adipose tissue (MAT) and corresponding lymph nodes (MLN) areharvested, and luminal and mucosal contents of each intestinal segmentseparated. Quantification of bacterial DNA is performed by isolatinggenomic DNA from blood, MAT, MLN or intestine (contents and mucosa). Allbacterial DNA is quantified by quantitative real-time PCR targetingconserved regions of the 16S rRNA gene, with bacterial DNA as standardtemplate for absolute quantification.

In order to assess barrier permeability, the expression of occludin andzonula occludens-1 (ZO-1) tight-junction proteins are assessed. Jejunumsegments are immediately removed, washed with PBS, mounted in embeddingmedium, and stored at −80° C. until use. Cryosections (5 mm) are fixedin acetone at −20° C. for 5 min for occludin and in ethanol for 30 minat room temperature and in acetone at −20° C. for 5 min for ZO-1.Non-specific background is blocked by incubation with 10% bovine serumalbumin (BSA) in Tris-buffered saline (TBS) and 0.3% Triton X-100 (30min at room temperature).

Sections are incubated with rabbit anti-occludin or rabbit anti-ZO-1(1:400 for ZO-1 and 1:100 for occludin staining) for 2 h. Sections arewashed three times for 10 min in TBS and probed with goat anti-rabbitfluorescein isothiocyante (FITC)-conjugated antibodies (1:50). Slidesare washed three times for 10 min in TBS and mounted in mounting medium.Sections are visualized on a fluorescence microscope. As a control,slides are incubated with serial dilutions of the primary antibody tosignal extinction. Two negative controls are used: slides incubated withirrelevant antibody or without primary antibody. All the stainings areperformed in duplicate in non-serial distant sections, and analyzed in adouble-blind manner by two different investigators.

The results show that HMS/HMO improve gut barrier function and reducethe metabolic inflammation and insulin resistance associated withobesity by increasing release of gut peptides, such as glucagon-likepeptide-1 and -2 (GLP-1 and -2).

Example 2—Treating Obesity Induced Diabetes

Six-week-old ob/ob mice (120 mice) on C57BL/6 background are housed in acontrolled environment (12 h daylight cycle) in groups of 2 mice/cage,and kept with free access to food and drinking water. The mice areseparated into 12 groups of 10 mice, one control group and 11 treatmentgroups. One group is fed a control diet, and the 11 treatment groupseach receive a control diets containing one of the following HMS/HMO (20g/kg of diet) for five weeks: a) sialic acid, b) L-fucose, c) 2′-FL, d)3-FL, e) 3′-SL, f) 6′-SL, g) LNT, h) LNnT, i) LNFP-I, j) DSLNT, and k) acombination of these saccharides. Fresh food is given daily.

Experiments to show impact of HMS/HMO on glucose tolerance, intestinalpermeability plasma LPS, cytokines and gut hormones, caecal microbiotaprofile and bacterial translocation are performed as described underExample 1.

Example 3—Human Trial in Overweight and Obese Children

A total of 60 male and female patients, enrolled to a childhood obesitytreatment program, are recruited to participate in the study. Patientsare randomized into three groups, each of 20 patients, with 2 groupsreceiving different investigational products and one group receiving aplacebo product for 8 weeks. The investigational products contain 4.5grams of either 2′-FL alone or a combination of 2′-FL and LNnT while theplacebo product contains 4.5 grams glucose. All products are in powderform in a unit dosage container.

The patients are eligible to participate if: they are between 5 and 10years of age, have a BMI SDS of ≥2.0 and are enrolled in the childhoodobesity treatment program at the Children's Obesity Clinic. Allrecruited patients and their representatives are able and willing tounderstand and comply with the study procedures. Patients are excludedif: they have participated in a clinical study one month prior to thescreening visit and throughout the study; have any gastrointestinaldisease(s) that may cause symptoms or may interfere with the trialoutcome; have other severe disease(s) such as malignancy, kidney diseaseor neurological disease; have psychiatric disease; have used highlydosed probiotic supplements (yoghurt allowed) 3 months prior toscreening and throughout the study; have consumed antibiotic drugs 3months prior to screening and throughout the study; and consume on aregular basis medication that might interfere with symptom evaluation 2weeks prior to screening and throughout the study.

At the initial visit (screening) patients and their representatives aregiven both oral and written information about the study; the childrenare asked for informed assent and their representatives to sign aninformed consent form.

Eligibility criteria are checked and for children who are enrolled tothe study, medical history and concomitant medication are registered. Aphysical examination is done and pubertal staging is determined. Bloodpressure, pulse rate, height and bodyweight are measured, and bodycomposition is determined by a DXA (dual energy x-rayabsorptiometry)-scan and bioimpedance. BMI SDS is calculated, waist andhip circumferences are measured and food intake is registered. Fastingblood samples are collected for safety and biomarker studies and forbiobanking.

The serum from the blood samples is transferred to cryotubes and storedat −80° C. The following biomarkers are measured; Lipopolysaccharides(LPS), hsCRP, free fatty acids, total cholesterol, HDL, LDL, HbA1c,glucose, insulin, triglycerides, TNF-α, IL-1β, IL-6, IL-8, IL-10, GLP-1,GLP-2, Adiponectin, and Zonulin.

Equipment for collecting faecal samples is distributed. The faecalsamples are stored at −80° C. until analysis. Microbiological analysisis performed on the faecal samples using 16S rRNA gene sequencing.

The Rome III Diagnostic Questionnaire for Paediatric Functional GIDisorders (QPFG) is completed on site by the participating child'srepresentative(s), and the Bristol Stool Form Scales (BSFS) isdistributed to the participant's representative(s) with instructions toassess the stool consistency at each faecal sampling point using theBSFS.

At the second visit (randomization), patients and their representativesare asked about adverse events, faecal samples are collected andequipment for collection of new samples is distributed. BSFS iscollected and new BSFS is distributed. Study products are distributedtogether with a compliance form (diary). Patients and theirrepresentatives are reminded to follow the healthy dietary habits.

The study runs for 8 weeks with the patients consuming either a placeboor one of two investigational products daily. Patients are instructed toconsume the products in the morning with breakfast. Compliance ismonitored via a compliance form (diary) to be filled in daily.

Four weeks after commencement there is an intermediate check. Patientsand their representatives are asked about adverse events and any changesin the patient's usual medication. Fecal samples are collected andequipment for collection of new samples is distributed. Blood pressure,pulse rate, waist and hip circumference, height and bodyweight aremeasured and BMI SDS calculated. The QPFG questionnaire is completed onsite by the participating child's representative. The BSFS is collectedand new BSFS is distributed to the participant's representative(s) withinstructions to assess the stool consistency at each faecal samplingpoint using the BSFS. Patients and their representatives are reminded tofollow the healthy dietary habits.

At the end of intervention (8 weeks), each patient has a visit with themedical team. Patients and their representatives are asked about adverseevents and any changes in the patient's usual medication. Study productsand compliance forms are collected to check compliance. BSFS and faecalsamples are collected and equipment for collection of new samples isdistributed. A physical examination is done and pubertal staging isdetermined. Blood pressure, pulse rate, height and bodyweight aremeasured, and body composition is determined by a DXA (dual energy x-rayabsorptiometry)-scan and bioimpedance. BMI SDS is calculated, waist andhip circumferences measured and food intake registered. Fasting bloodsamples are collected for safety and biomarker studies and forbiobanking, and equipment for collecting faecal samples is distributed.The QPFG questionnaire is completed on site by the participating child'srepresentative(s).

To examine potential long term effects of the intervention, anun-blinded follow-up period follows with a visit 8 weeks after end ofintervention. A physical examination is done and pubertal staging isdetermined. Blood pressure, pulse rate, height and bodyweight aremeasured, and body composition is determined by a DXA (dual energy x-rayabsorptiometry)-scan and bioimpedance. BMI SDS is calculated, waist andhip circumferences measured and food intake registered. Fasting bloodsamples are collected for safety and biomarker studies and forbiobanking. Fecal samples are collected. The intervention contributes toa normal body composition, and the patients given the investigationalproducts show a greater reduction of body fat, body weight and BMI SDSas compared to the placebo group. The blood biomarker analysis indicatesthat the patients given the investigational products have increasedlevels of GLP-1 and GLP-2, reduced levels of metabolic endotoxemia andinflammatory markers and reduced gut permeability indicating an improvedmucosal barrier compared to the placebo. The faecal analysis indicatesthat the patients given the investigational products have reducedbacterial dysbiosis and a higher level of bifidobacteria compared to theplacebo, particularly Bifidobacteria adolescentis.

Example 4—Nutritional Composition

A ready to feed nutritional composition is prepared from water,maltodextrin, milk protein concentrate, Sucromalt, glycerine, cocoapowder, soy protein isolate, fructose, high oleic safflower oil, soyoil, canola oil, plant sterol esters, HMSs/HMOs, soy lecithin, magnesiumchloride, calcium phosphate, carrageenan, sodium ascorbate, potassiumcitrate, sodium phosphate, calcium citrate, choline chloride, potassiumchloride, sodium citrate, magnesium oxide, taurine, L-carnitine,alpha-tocopheryl acetate, zinc sulphate, ferrous sulphate, niacinamide,calcium pantothenate, vitamin A palmitate, citric acid, manganesesulphate, pyridoxine hydrochloride, vitamin D3, copper sulphate,thiamine mononitrate, riboflavin, beta carotene, folic acid, biotin,potassium iodide, chromium chloride, sodium selenate, sodium molybdate,phytonadione, vitamin B12.

The composition has an energy density of 0.8 kcal/ml with an energydistribution (% of kcal) as follows: protein: 20%, carbohydrate: 48%,fat: 32%.

Example 5—Tablet Composition

A tablet is prepared from HMS/HMO, hydroxypropyl methylcellulose, sodiumalginate, gum, microcrystalline cellulose, colloidal silicon dioxide,and magnesium stearate. All raw materials except the magnesium stearateare placed into a high shear granulator and premixed. Water is sprayedonto the premix while continuing to mix at 300 rpm. The granulate istransferred to a fluidized bed drier and dried at 75° C. The driedpowder is sieved and sized using a mill. The resulting powder is thenlubricated with magnesium stearate and pressed into tablets. The tabletseach contain 325 mg of HMS/HMO. The tablets each have a weight of 750mg.

Example 6—Capsule Composition

A capsule is prepared by filling about 1 g of HMS/HMO into a 000gelatine capsule using a filing machine. The capsules are then closed.The HMS/HMO are in free flowing, powder form.

What is claimed is:
 1. A method comprising: determining a treatmentgroup comprising obese non-infant humans, formulating a compositioncomprising an effective amount of one or more synthetic fucosylatedhuman milk oligosaccharides (HMO) selected from 2′-fucosyllactose(2′-FL), 3-fucosyllactose (3-FL), difucosyllactose (DFL), andlacto-N-fucopentaose I (LNFP-I), that are effective for: increasing inthe gastrointestinal microbiota of a non-infant human during a treatmentperiod, the relative abundance of Bifidobacterium adolescentis; andreducing a precursor condition for a metabolic disorder associated withdevelopment of one or more of obesity-induced pre-diabetes and type 2diabetes, the precursor condition selected from gut permeability,metabolic endotoxemia, low-grade metabolic inflammation, and body fatpercentage; reducing the precursor condition in at least one non-infanthuman in the treatment group by providing the composition to the atleast one non-infant human during the treatment period.
 2. The method ofclaim 1, wherein the reduced precursor condition for the metabolicdisorder associated with development of the one or more ofobesity-induced pre-diabetes and type 2 diabetes is gut permeability. 3.The method of claim 1, wherein the reduced precursor condition for themetabolic disorder associated with development of the one or more ofobesity-induced pre-diabetes and type 2 diabetes is body fat percentage.4. The method of claim 1, further comprising increasing, in thegastrointestinal tract of the non-infant human, a level of glucagon-likepeptide selected from GLP-1 and/or GLP-2 relative to the level of theselected glucagon-like peptide prior to the treatment period.
 5. Themethod of claim 1, wherein: the treatment period comprises an initialtreatment phase and a maintenance phase; the effective amount of theselected one or more HMOs is from about 2.5 g to about 7.5 g dailyduring the initial treatment phase; and the effective amount of theselected one or more HMOs is from about 1 g to about 2.5 g daily duringthe maintenance phase.
 6. The method of claim 1, further comprisingformulating the composition in a unit dosage form.
 7. The methodaccording to claim 1, wherein the obese non-infant human is aprepubescent child.
 8. A method comprising: determining a treatmentgroup comprising obese non-infant humans; formulating a compositioncomprising one or more synthetic non-fucosylated human milkoligosaccharides (HMOs) selected from lacto-N-tetraose (LNT),lacto-N-neotetraose (LNnT), that are effective for: increasing in thegastrointestinal microbiota of a non-infant human during a treatmentperiod, the relative abundance of Bifidobacterium adolescentis andreducing a precursor condition for a metabolic disorder associated withdevelopment of one or more of obesity-induced pre-diabetes and type 2diabetes, the precursor condition selected from gut permeability,metabolic endotoxemia, low-grade metabolic inflammation, and body fatpercentage; and reducing the precursor condition in at least onenon-infant human in the treatment group by providing the composition tothe at least one non-infant human during the treatment period.
 9. Themethod of claim 8, wherein the reduced precursor condition for themetabolic disorder associated with development of the one or more ofobesity-induced pre-diabetes and type 2 diabetes is gut permeability.10. The method of claim 8, wherein the reduced precursor condition forthe metabolic disorder associated with development of the one or more ofobesity-induced pre-diabetes and type 2 diabetes is body fat percentage.11. The method of claim 8, further comprising increasing, in thegastrointestinal tract of the non-infant human, a level of glucagon-likepeptide selected from GLP-1 and/or GLP-2 relative to the level of theselected glucagon-like peptide prior to the treatment period.
 12. Themethod of claim 8, further comprising formulating the composition in aunit dosage form.
 13. The method according to claim 8, wherein the obesenon-infant human is a prepubescent child.
 14. The method of claim 8,wherein: the treatment period comprises an initial treatment phase and amaintenance phase; the effective amount of the selected one or more HMOsis from about 2.5 g to about 7.5 g daily during the initial treatmentphase; and the effective amount of the selected one or more HMOs is fromabout 1 g to about 2.5 g daily during the maintenance phase.
 15. Amethod comprising: determining a treatment group comprising obesenon-infant humans; formulating a composition comprising an effectiveamount of two or more synthetic neutral human milk oligosaccharides(HMOs) selected from 2′-fucosyllactose (2′FL), 3-fucosyllactose (3-FL),difucosyllactose (DFL), lacto-N-fucopentaose I (LNFP-I),lacto-N-tetraose (LNT), and lacto-N-neotetraose (LNnT), wherein theselected HMOs are effective for: increasing in the gastrointestinalmicrobiota of a non-infant human during a treatment period, the relativeabundance of Bifidobacterium adolescentis, and reducing in thenon-infant human during the treatment period, a precursor condition fora metabolic disorder associated with development of one or more ofobesity-induced pre-diabetes and type 2 diabetes, the precursorcondition selected from gut permeability, metabolic endotoxemia,low-grade metabolic inflammation, and body fat percentage; reducing theprecursor condition in at least one non-infant human in the treatmentgroup by providing the composition to the at least one non-infant humanduring the treatment period.
 16. The method of claim 15, wherein thereduced precursor condition for the metabolic disorder associated withdevelopment of the one or more of obesity-induced pre-diabetes and type2 diabetes is gut permeability.
 17. The method of claim 15, wherein thereduced precursor condition for the metabolic disorder associated withdevelopment of the one or more of obesity-induced pre-diabetes and type2 diabetes is body fat percentage.
 18. The method of claim 15, furthercomprising increasing, in the gastrointestinal tract of the non-infanthuman, a level of glucagon-like peptide selected from GLP-1 and/or GLP-2relative to the level of the selected glucagon-like peptide prior to thetreatment period.
 19. The method of claim 15, further comprisingformulating the composition in a unit dosage form.
 20. The method ofclaim 15, wherein: the treatment period comprises an initial treatmentphase and a maintenance phase; the effective amount of the selected HMOmixture is from about 2.5 g to about 7.5 g daily during the initialtreatment phase; and the effective amount of the selected HMO mixture isfrom about 1 g to about 2.5 g daily during the maintenance phase.